The genome-scale metabolic network analysis of Zymomonas mobilis ZM4 explains physiological features and suggests ethanol and succinic acid production strategies

Abstract

BackgroundZymomonas mobilis ZM4 is a Gram-negative bacterium that can efficiently produce ethanol from various carbon substrates, including glucose, fructose, and sucrose, via the Entner-Doudoroff pathway. However, systems metabolic engineering is required to further enhance its metabolic performance for industrial application. As an important step towards this goal, the genome-scale metabolic model of Z. mobilis is required to systematically analyze in silico the metabolic characteristics of this bacterium under a wide range of genotypic and environmental conditions.ResultsThe genome-scale metabolic model of Z. mobilis ZM4, ZmoMBEL601, was reconstructed based on its annotated genes, literature, physiological and biochemical databases. The metabolic model comprises 579 metabolites and 601 metabolic reactions (571 biochemical conversion and 30 transport reactions), built upon extensive search of existing knowledge. Physiological features of Z. mobilis were then examined using constraints-based flux analysis in detail as follows. First, the physiological changes of Z. mobilis as it shifts from anaerobic to aerobic environments (i.e. aerobic shift) were investigated. Then the intensities of flux-sum, which is the cluster of either all ingoing or outgoing fluxes through a metabolite, and the maximum in silico yields of ethanol for Z. mobilis and Escherichia coli were compared and analyzed. Furthermore, the substrate utilization range of Z. mobilis was expanded to include pentose sugar metabolism by introducing metabolic pathways to allow Z. mobilis to utilize pentose sugars. Finally, double gene knock-out simulations were performed to design a strategy for efficiently producing succinic acid as another example of application of the genome-scale metabolic model of Z. mobilis.ConclusionThe genome-scale metabolic model reconstructed in this study was able to successfully represent the metabolic characteristics of Z. mobilis under various conditions as validated by experiments and literature information. This reconstructed metabolic model will allow better understanding of Z. mobilis metabolism and consequently designing metabolic engineering strategies for various biotechnological applications.

N2gas is an effective fertilizer for bioethanol production byZymomonas mobilis

“…We used 13 C-labeling studies to determine whether intermediary metabolism changed in response to growth with N 2 . Metabolic fluxes were estimated from measurements of glucose uptake and ethanol production, the ZM4 biomass composition (15), and amino acid labeling patterns generated by metabolic pathways that processed the 13 C-labeled glucose (Table S3). The resulting flux distributions, normalized to the glucose uptake rate, were similar regardless of the nitrogen source, except for biosynthetic fluxes, which were twice as high during growth with NH 4 + compared with growth with N 2 ( Fig.…”

“…Metabolic fluxes were estimated from the glucose uptake rate, the ethanol production rate, the biomass composition for Z. mobilis ZM4 (15), and mass isotopomer distributions from parallel labeling experiments (i.e., using 100% [1-13 C]glucose in one experiment and a mixture of 80% unlabeled and 20% uniformly labeled glucose in the other). Mass isotopomer distributions were corrected for natural isotopic abundances using IsoCor software (31).…”

Comparative Analysis of Biochemical Network Reconstructions

“…Two genome-scale reconstructions of Zymomonas mobilis by Lee et al, (2010) having 615 metabolites and 600 reactions, and Widiastuti et al, (2011) having 773 metabolites and 747 reactions were compared.…”

Droplet Microfluidic Technique for the Study of Fermentation

“…The first approach concentrates on searching for strains displaying the ability to metabolize other sugars, e.g., cellulose [10] or xylose [11]. The second approach focuses on inducing mutagenesis resulting in, e.g., ability to simultaneously use xylose and glucose by Z. mobilis [12] or an increase of tolerance for the presence of metabolite products [13]. The widespread interest in the use of Z. mobilis in production of biofuels motivates the search for precise and easy-to-use assays.…”